Alpha 2 macroglobulin gene and their expression in response to GFP tagged Vibrio parahaemolyticus and WSSV pathogens in Indian white shrimp Fenneropenaeus indicus

Alpha 2 macroglobulin (α2M) is a non-specific protease inhibitor involved in host defence mechanism in invertebrates. By c-DNA cloning and sequencing, α2M gene was identified from the haemocytes of Indian white shrimp Fenneropenaeus indicus with an open reading frame of 4599 bp encoding 1532 amino acids with a signal sequence of 26 amino acids. The calculated molecular mass of the mature protein is 170.6 kDa with an estimated pI of 6.13. The F. indicus α2M (Fi-α2M) sequence contains putative conserved functional domains including a GCGEQNM thioester region and a receptor-binding domain which are present in other invertebrate and vertebrate α2Ms. Sequence comparison and phylogenetic analysis revealed that Fi-α2M displays the highest similarity with Litopenaeus vannamei (96%). QRT-PCR analysis of challenge experiments by Vibrio harveyi, GFP tagged Vibrio parahaemolyticus (GFP-Vp) and WSSV infection of F. indicus significantly increased the α2M mRNA transcript up to 24 h in the haemocytes. In addition, confocal laser scanning microscopy (CLSM) confirms that the GFP-Vp treated F. indicus have high proliferation of GFP-Vp in haemolymph and intestine at 24 h. These results suggest that α2M is an important molecule in shrimp immune system.